University of Khartoum

Oligopeptidase B, a Potential Drug Target for Infectious Diseases

Oligopeptidase B, a Potential Drug Target for Infectious Diseases

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Title: Oligopeptidase B, a Potential Drug Target for Infectious Diseases
Author: Mohamed, MS; Nakajima, Y; Iwata, N
Abstract: The oligopeptidase B (OPB; EC 3.4.21.83) subfamily represents one of two branches in the prolyl oligopeptidase family of serine peptidases (the S9 family, according to the nomenclature of Barrett and Rawlings) which cleave peptides composed of less than 30 amino acids at the carboxyl side of arginine or lysine residue. In this study, we have amplified oligopeptidase B genes by PCR from 2 gram-negative bacteria Stenotrophomonas maltophilia (Stm) and Serratia marcescens (Sem) and a gram-positive Rhodococcus erythropolis (Re), in addition to E. coli (EC) OPB gene for comparison. The PCR products were cloned into pET expression vectors (Novagen) and the nucleotide sequences of OPB genes were confirmed. E. coli BL21 DE3 was used as a host for expression and the expressed enzymes were extracted by sonication and purified by a metal-chelating column charged with Ni2+ and ion-exchange column chromatography. Kinetic parameters, substrate specificities and some physicochemical properties of the recombinant proteins were determined. Strong substrate inhibition as a common feature of Stm, Ec and ReOPBs with substrates containing double Arg residues was observed, while SemOPB was found to be the only enzyme that was not inhibited by these substrates. This study highlights the importance of SeOPB for studying the mechanism of substrate inhibition, a phenomenon which is crucial in the process of drug discovery. Preliminary crystallization screening of the enzymes was carried out and some crystals of ReOPB and StmOPB were obtained. These crystals would be used for structural determination and structural-based drug design to identify inhibitors as potential antimicrobial agents
URI: http://khartoumspace.uofk.edu/handle/123456789/19594
Date: 2015


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